The line of code you asked about basically says if $columns[1] is true (has any value Perl evaluates as true) and contains a string that begins with "HLA-A" then take the following actions. I included the first "does it have a true value" check because I assumed use warnings; would end up complaining for any line that didn't have an element at index 1 in $columns. I didn't actually try it without it, but I just assumed that would happen for at least the all "---" lines.

As for the code changes you requested:

#!/usr/bin/perl
use warnings;
use strict;

open(my $in_fh, '<', 'input.txt') or die $!;
my $output;

my %seen_lines;
while (<$in_fh>)
{
    chomp;
    my @columns = split;
    
    if ($columns[1] and $columns[1] =~ /^HLA-A/)
    {
        my $HLA_Peptide = $columns[1] . $columns[2];
        $output .= "$_\n" if (!exists $seen_lines{$HLA_Peptide});
        $seen_lines{$HLA_Peptide} = 1;
    }
    else
    {
        $output .= "$_\n";
    }
}
close $in_fh;

print $output;
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EDIT: I did just try it without that first check and I was correct, it does throw warnings without it. There may be a better way to avoid that warning (it does occur to me that false values like "0" or an empty string would be evaluated as such), but I use this trick a lot in an attempt to appease use warnings; or "-w". I wonder if there is something like exists which I use a lot for hashes only meant for use to check if an array element exists?